The antitumor toxin CD437 is a direct inhibitor of DNA polymerase α

Abstract

CD437 is a retinoid-like small molecule that selectively induces apoptosis in cancer but not normal cells through an unknown mechanism. We used a forward genetic strategy to discover mutations in POLA1 that coincide with CD437 resistance (POLA1R). Introduction of one of these mutations into cancer cells by CRISPR/Cas9 genome editing conferred CD437 resistance demonstrating causality. POLA1 encodes DNA polymerase α, the enzyme responsible for initiating DNA synthesis during the S phase of the cell cycle. CD437 inhibits DNA replication in cells and recombinant POLA1 activity in vitro. Both effects are abrogated by mutations associated with POLA1R. In addition, we detected an increase in the total fluorescence intensity and anisotropy of CD437 in the presence of increasing concentrations of POLA1 consistent with a direct binding interaction. The discovery of POLA1 as the direct anti-cancer target for CD437 has the potential to catalyze its development into an anti-cancer therapeutic.

Physiological Tolerance to ssDNA Enables Strand Uncoupling during DNA Replication

“…A recent study identified the adamantyl retinoid CD437 as a direct allosteric inhibitor of POLA1 (Han et al, 2016). To test whether CD437 can mimic the effect of POLA1 depletion, we first exposed U2OS cells to CD437 and measured CB-RPA as before.…”

“…In comparison to CD437, other commonly used inhibitors of DNA synthesis, such as hydroxyurea (HU) or aphidicolin (APH) that prevent the activity of all replicative polymerases ( Figure S2B), caused only a mild or undetectable increase in CB-RPA (Figure 2D). To confirm that POLA1 inhibition was responsible for the appearance of ssDNA, we analyzed the response to ST1926 and CD437 in CD437-resistant cells (Han et al, 2016). These cells express a mutated POLA1 that conserves its enzymatic activity but is resistant to CD437 in vitro.…”

Discovery of a molecular glue promoting CDK12-DDB1 interaction to trigger Cyclin K degradation

“…To identify any target protein(s) destabilized by HQ461, we performed gain-of-function genetic screening in the colorectal cancer cell line HCT-116. HCT-116 cells are defective in mismatch repair, and therefore harbor a high rate of random point mutations 20,21 . Previous studies suggest that the identification of recurrent mutations present in multiple independent drug-resistant clones may reveal the direct drug target 8,21 .…”

“…HCT-116 cells are defective in mismatch repair, and therefore harbor a high rate of random point mutations 20,21 . Previous studies suggest that the identification of recurrent mutations present in multiple independent drug-resistant clones may reveal the direct drug target 8,21 . To ensure the isolation of independent HQ461-resistant clones, we derived 10 clonal isolates from HCT-116 that were initially sensitive to HQ461 (HQ461 S ).…”

Repurposing bioactive compounds for treating multidrug-resistant pathogens

“…The lattermost category included DMAQ-B1, an insulin mimetic purified from Pseudomassaria sp. [24]; CD437, an activator of retinoic acid receptor-β and -γ [25]; oxaliplatin and carboplatin, FDA approved platinum complexes for the treatment of cancer [26,27]; and PSB-069, a nucleoside triphosphate diphosphohydrolase inhibitor [28] (Fig. 1a).…”